Hematopoietic factors represented by erythropoietin (EPO) and granulocyte-colony stimulating factor (G-CSF) have already been developed as pharmaceutical agents, and have been utilized for the treatment of various diseases. Cytokines, such as interferon, and hormones, including insulin and growth hormone, are also commercially available as pharmaceutical agents. Most of the biologically active proteins used as pharmaceutical agents have been produced by genetic engineering techniques. Low molecular weight compounds are being screened as the next-generation drugs replacing biologically active proteins. For instance, if it was possible to find low molecular weight compounds having the same activities as biologically active proteins, such compounds will be useful as pharmaceutical agents, since they may be orally administered. Therefore, there is a need to develop efficient methods for screening low molecular weight compounds that have biological activities similar to naturally existing ligands.
Several screening methods are already known. For example, when screening for a ligand of a receptor whose amino acid sequence and function are known, there is a method in which a chimeric receptor comprising the intracellular domain of a receptor with a known function and the extracellular domain of a receptor of interest is prepared, and the chimeric protein is used to screen for a ligand of the receptor of interest (Ishizaka-Ikeda, E., Pro. Natl. Acad. Sci. USA (1993) 90, p123-127; U.S. Pat. No. 4,859,609 and U.S. Pat. No. 5,030,576). Using the insulin receptor as the extracellular domain and the EGF receptor as the intracellular domain, these US patents describe a method that assays in a cell-free system chimeric receptor phosphorylation induced by the binding of a ligand. A method using a chimeric receptor that consists of the extracellular domain of the EGF receptor and the intracellular domain of the EPO receptor, has also been reported (WO94-29458). These approaches may also be applicable to the screening of ligands for an orphan receptor, whose natural ligand is not yet known.
However, in each of these screening methods, only one kind of receptor is used as the receptor of interest, and one screening can only detect the effect a test sample has on that single receptor. Therefore, in order to assay the effect on several receptors, the same number of screenings as that of receptors of interest is required. Also, these approaches are inefficient for a preliminary screening of a vast number of test samples that have different structures and whose activities are unconfirmed, since activities of most ligands cannot be detected. It is, therefore, necessary to develop a method that can efficiently and rapidly screen a vast number of test samples.